The goal of the proposed research program is to establish the mechanism(s) for reversible myocardial dysfunction following the systemic release of tumor necrosis factor-alpha (TNFalpha) as would occur during an episode of bacteremia or endo-toxemia. The unifying hypothesis for this work is that TNFalpha interacts directly with the myocardium, so that within 24 hours, there are derangements in intracellular physiology that either compromise contractile function or permit myocardial injury by secondary mediators. In the first series of experiments, chronically instrumented conscious rabbits will be studied to determine whether secondary cellular and/or humoral mediators are necessary for myocardial dysfunction induced by recombinant human (rh)TNFalpha. In these experiments, the infiltration of the left ventricular (LV) myocardium by neutrophils (PMNL) and monocytes will be monitored in animals in which either PMNL and monocytes have been depleted or the CD-18 adhesiveness molecule on PMNL and monocytes has been blocked with antibody. To determine whether prostaglandins, leukotrienes, platelet activating factor, or active O2-derived species are necessary for myocardial dysfunction, experiments will be conducted in which animals are pretreated with a cylooxygenase inhibitor, leukotriene receptor antagonist, platelet activating factor antagonist, or a scavenger of active O2 metabolites. The second series of experiments will characterize the myocardial dysfunction induced by rhTNFalpha in unanesthetized dogs by examining relationships between mechanical energy expenditure, myocardial blood flow, and myocardial O2 consumption (MVO2). These experiments will test the hypothesis that myocardial dysfunction after rhTNFalpha is the result of derangements that alter O2 availability and/or utilization. The following parameters will be examined during the period of myocardial dysfunction and during recovery: 1) myocardial blood flow and distribution, 2) myocardial O2 extraction and utilization, 3) the efficiency of O2 utilization for mechanical energy expenditure, and 4) the fractional utilization of O2 for non-contractile function. The third series of experiments will investigate whether exposure of the myocardium to rhTNFalpha induces alterations in gene expression and protein synthesis that might potentially compromise contractile function. To determine whether altered protein synthesis is the direct result of myocardial exposure to rhTNFalpha or whether secondary mediators are necessary, mRNA's and proteins will be examined both in myocardial tissue harvested from in vivo experiments and in myocardial cells grown in culture. Whether there is up- or down- regulation of the overall levels of mRNA and protein synthesis after exposure to rhTNFalpha will be examined. Alterations in the expression of specific proteins related to myocardial contractility (e.g. heavy-chain myosin subtypes), myocardial energetics (e.g. creatine kinase), or the cellular response to injury (e.g. stress proteins, catalase) will also be studied. These studies should provide important insights into the mechanism for temporary myocardial dysfunction in critical illness induced by sepsis. The data generated will be of considerable interest for two reasons: 1) impaired myocardial performance is probably an important factor contributing to morbidity and mortality in septic shock; and 2) the heart represents a unique organ for studying the relationship between function (i.e., mechanical work) and metabolism (MVO2, protein synthesis).